Thermodynamic And Kinetic Studies Of Protein Structure A

本实验室致力于研究蛋白质序列、结构与蛋白质折叠和酶反应机制之间的关系。(I)不溶性蛋白质的二级结构。蛋白质在溶液中的结构很容易通过对其圆二色谱(CD)的数学分析来估计。与普恩病毒疾病有关的蛋白质是不可溶的，必须作为薄膜进行研究。在这种情况下，通常对CD数据进行量化和分析所必需的蛋白质浓度无法确定。我们已经展示了一种不依赖于浓度的结构确定方法，该方法包括测量和解卷积蛋白质的“g因子”谱，即样品CD与吸收光谱的比率。(Ii)胱硫醚β合成酶的结构(与伊迪丝·威尔逊·迈尔斯博士及同事，LBG)我们正在研究色氨酸合成酶α2-β2复合体的构象状态及其与蛋白质的酶机制的相关性。(Iii)哺乳动物硫酸盐转移酶的结构。(与D.马歇尔博士和W.B.Jakoby博士，LBM)来自大鼠肝脏的苯酚硫代转移酶的酶活性已被证明是通过生理浓度的谷胱甘肽可逆地氧化/还原保守的特定半胱氨酸残基来调节的。半胱氨酸残基66的氧化通过非常紧密的底物抑制来抑制酶的生理活性。这一机制在氧化应激对肝脏的条件下可能是重要的。(Iv)轮状病毒蛋白NSP2的结构(与Peter Schuck博士，ORS，OD和Z.Taraporewala博士，LID，NIAID)该蛋白是病毒复制装置的组成部分，与RNA和核苷酸相互作用。我们正在研究它的结构和与配体相互作用引起的变化。

This laboratory is interested in the relationship among protein sequence, structure and the mechanisms of protein folding and enzymic reactions. (i) Secondary Structure of Insoluble Proteins. The structure of a protein in solution is readily estimated by mathematical analysis of its circular dichroism (CD)spectrum. Proteins implicated in prion diseases are insoluble and must be studied as thin films. Under these circumstances, protein concentration, which is usually necessary for quantification and analysis of CD data cannot be determined. We have shown a concentration independent method of structure determination, which involves measurement and deconvolution of the protein's "g-factor" spectrum, which is the ratio of a samples CD and absorbance spectra. (ii) Structure of Cystathionine beta Synthase (with Dr. Edith Wilson Miles and coworkers, LBG) We are investigating the conformational states of the tryptophan synthase alpha2-beta2 complex and their relevance in the enzymatic mechanism of the protein. (iii) Structure of Mammalian Sulphotransferases. (with Dr. D. Marshall and Dr. W.B. Jakoby, LBM) The enzymatic activity of a phenol sulphotransferase, from rat liver, has been shown to be regulated by reversible oxidation/reduction of a conserved specific cysteine residue by physiological concentrations of glutathione. Oxdation of cysteine residue 66 inhibits the physiological activity of the enzyme by very tight substrate inhibition. This mechanism may be important under conditions of oxidative stress to the liver. (iv) Structure of Rotavirus protein NSP2 (with Dr. Peter Schuck, ORS, OD and Dr. Z. Taraporewala, LID, NIAID) This protein is a component of the virus' replication apparatus, interacting with RNA and nucleotides. We are studying its structure and the changes induced by interaction with ligands.